November 2011 Funded Microgrants

November 2011 Funded Microgrants

Clinical and Radiological Investigation of Cerebellar Atrophy in Childhood

Children with ataxia due to cerebellar atrophy face a number of challenges in their daily lives. Many of these disorders are due to genetic causes, and accurate diagnosis is important for better management and counseling of these families. We would like to establish a diagnostic algorithm that will enable early and accurate diagnosis of childhood cerebellar atrophy.

Long QT Syndrome with Induced Pluripotent Stem Cells

The goal of this study is to better understand how heart cells of patients diagnosed with LQT Syndrome (LQTS) respond differently to medications, environments and circumstances. This syndrome is a rare heart condition that causes the heart to beat irregularly, also known as arrhythmia, and lead to serious consequences such as dizziness, fainting or cardiac arrest and sudden death.

The proposed study is designed by a pediatric cardiologist specializing in cardiac arrhythmias and a molecular cardiologist. To study the cause of heart beat irregularity, they will develop a model that mimics LQTS patients' heart cells by converting skin cells of willing participants with LQTS to 'induced pluripotent stem cells" (iPSCs), which will then be engineered to transform into heart cells under laboratory settings. Engineered heart cells from LQTS patients will be compared with iPSC-derived heart cells of healthy control participants without LQTS to better understand abnormalities associated with the syndrome. If this cell model can be successfully developed, the investigators will be able to gain important insights on potential triggers that can lead to deadly cardiac arrhythmias in LQTS patients. This understanding may help to determine which medications or other health care interventions would be best for preventing cardiac arrhythmias in patients with LQTS.

Use of a Simple Breath Test to Examine Kuvan® Responsiveness in Children with PKU

Phenylketonuria (PKU) is an autosomal recessive disorder caused by deficiency of an enzyme in the liver called phenylalanine hydroxylase (PAH). Incidence of PKU is reported at 1 in 10,000 babies. PAH deficiency leads to increased levels of phenylalanine (an amino acid) in plasma. The disease causes intellectual disability unless the affected child is maintained on a strict low phenylalanine diet. A new drug called Kuvan® (Sapropterin dihydrochloride), a synthetic form of the tetrahydrobiopterin (BH4) which is used by the PAH enzyme has shown promise in decreasing the levels of plasma phenylalanine. But the response to this drug varies among the children. In some children there is a very small decrease in plasma phenylalanine concentrations. In some children even when there is no change in plasma phenylalanine the child seems to improve in cognitive functions. Thus, there is a need to develop a test that will measure whether the drug is actually responsive. The new stable isotope (a very safe form of isotope, widely used in medical research) test uses phenylalanine, with a 13C label. If the child is responding to Kuvan®, then there will be increased amounts of 13CO2 in the breath, as the 13C-Phenylalanine will be metabolized by the liver and the 13C released in breath as 13CO2. Thus, this simple breath test before and after the provision of Kuvan® will help in identifying children who respond to treatment and will help immensely in management of children with PKU. The current proposal is to perform pilot studies on two PKU children, who have been identified as good candidates for the drug Kuvan®.

Induced Pluripotent Stem Cell Models of Alagille Syndrome

Our objective is to develop cell-based models of Alagille syndrome (ALGS) using innovative induced pluripotent stem cell (iPSC) technology. ALGS is an inherited condition (with a frequency of 1 in 30,000), manifest by insufficient bile ducts in the liver. Liver disease starts in infancy and is associated with significant morbidity and mortality. We are unable to predict liver disease outcomes in ALGS and this poses a management challenge. The timing and necessity of interventions such as liver transplantation would be greatly assisted by a patient-specific model of liver disease. iPSCs are derived from skin cells and have the potential to differentiate into any cell type. Thus we can use a patient's own skin sample and develop a patient-specific disease model in order to understand their disease, predict outcomes and test new therapies. This has been successful with heart and liver cells and here we plan to develop a protocol to generate bile duct cells. A bile duct model of ALGS would have a direct applicability to personalizing medical care and also allow us to easily screen novel drugs.

Glucose is the main energy source for the normal brain function and is continuously delivered into the brain by a transporter protein called GLUT1. Individuals with deficiency of this transporter protein have seizures, small head size, coordination problems, speech delay and intellectual disability secondary to decreased glucose supply into brain. A high-fat, low-carbohydrate diet, called ketogenic diet, is the specific therapy for this disease. The diagnosis is confirmed by molecular genetic testing of the GLUT1 gene. We will review patients in the Division of Neurology to identify with various epilepsy types to investigate those individuals for the GLUT1 deficiency, a treatable metabolic epilepsy.

Results - I would like to thank the Rare Disease Foundation for their generous support of our project, funded in November 2011. We finalized our study and published our results recently (Hewson et al, Canadian Journal of Neurological Sciences, 2018). We reviewed charts for 85 patients and 18 of those had a genetic diagnosis including GLUT1 deficiency, pathogenic copy number variants, congenital disorder of glycosylation, neuronal ceroid lipofuscinosis type II, mitochondrial disorders, tuberous sclerosis, lissencephaly, and SCN1A-, SCN8A-, and STXBP1-associated epileptic encephalopathies. The prevalence of genetic diagnoses was 21% and prevalence of GLUT1 deficiency was 2.4% in our retrospective cohort study. Your support made this study possible. Thank you very much for the ongoing funding support.

Newborn Screening Outcomes for Inborn Errors of Metabolism

Newborn screening (NBS) is a public screening program where several medical conditions are screened for and if positive, children can be treated pre-symptomatically. We aim to compare the outcome of these children with the natural history of the conditions and with other NBS centers.

The Genetic Etiology of Aicardi Syndrome - Insights into a Developmental Disorder

Aicardi syndrome is a rare condition associated with severe developmental delay, brain malformation, seizures, and characteristic lesions on the retina or optic nerve. This disorder appears only to affect girls. Even though it was first reported over 40 years ago, the cause remains elusive. If we can understand what causes Aicardi syndrome, we may be able to provide more directed care for these girls. Also, understanding this condition may shed light onto early brain development, providing insight into many other disorders. We propose to study patients identified through our hospital to try to identify the gene involved in Aicardi Syndrome.

Delineation of Clinical Features of Individuals with Microduplication 22q11.2: A Questionnaire Study

The purpose of this study is to survey parents of individuals with 22q11.2 microduplication to determine the range of associated medical, cognitive and behavioural problems. Using an online parent questionnaire, we will collect information about family history, pregnancy, birth defects, developmental milestones, current academic and behavioural functioning, medical diagnoses and general health. We hope that collecting and summarizing this information will help guide patients and their health care professionals when making health care decisions.

Myoclonus is brief involuntary muscle twitching. If it is frequent, it may interfere with daily life of children and adolescents. When myoclonus is accompanied by abnormal electrical activity in brain, is called myoclonic epilepsy (ME). Progressive myoclonic epilepsy (PME) is the most severe form of ME. Patients have uncontrollable muscle twitches and a decline in cognitive ability because of nonresponsive seizures to anti-epileptic drugs due to underlying cause. More than 40 conditions can cause PME and some of them can be treatable with various supplements and vitamins. We will review Neurology Clinic charts and EEG's to identify individuals with ME and PME. We will generate a database for future studies to investigate individuals with ME and PME for treatable metabolic epilepsies.

Clinical Presentation of Mosaic Individuals

Approximately 90% of individuals with 45,X/46,XY mosaicism are normal males at birth, however, the remaining 10% may be male or female and have short stature, internal and/or external genital ambiguity, issues with puberty, or distinct physical features. The published literature on this population does not include long-term follow-up of these children and little information on the psychological concerns. We propose to review the hospital charts of individuals with 45,X/46,XY mosaicism in a population of 80 individuals. The goal of this study is to learn more about this unique condition with the hopes of better informing their medical and psychological management. In addition, parents faced with a prenatal diagnosis of 45,X/46,XY mosaicism can be better informed of the issues and concerns they can expect for their child.

Outcomes Study for Children with Non-Medullary Thyroid Carcinoma

Thyroid cancer is rare in children, however, it is increasing in frequency. Approaches to treatment vary widely among physicians and institutions. Because of this, it has been difficult to identify the most effective treatment regimens and the most appropriate tests to follow children after their cancer therapy. Five years ago, our hospital standardized its approach to treatment and follow-up of children with thyroid cancer. We will analyze the outcomes of this approach and, based on these results, we will revise the protocol with the goal of maximizing cure, while limiting complications and restricting testing to only that which is necessary.

Identification of a Genetic Cause for Recurrent Trisomy 21

Down Syndrome is a genetic condition in which patients present with a broad range of learning difficulties, as well as other health concerns such as congenital heart defects, and eye, hearing and digestive problems. Over 95% of affected patients have an additional chromosome 21 (trisomy 21) in their cells. This is usually a sporadic condition associated with an extra chromosome in the egg or sperm of a parent. The risk of having a child with Trisomy 21 increases with maternal age, and couples with an affected child are at slightly increased risk to have another affected child in subsequent pregnancies. We have identified a family with trisomy 21 in four close relatives in 3 generations. We believe they have a genetic condition that predisposes healthy individuals to have children with trisomy 21. Understanding the genetic cause will help to understand how the altered cell division that underlies trisomy 21 happens and may allow the identification of individuals who are at an increased risk.

Galactosialidosis and Emphysema: Is There Abnormal Elastogenesis in This Lysosomal Disease?

Lysosomal diseases are a group of rare diseases caused by failed processing in the cell's recycling centre-the lysosome-and resulting over-storage of unprocessed materials. Sometimes there are effects on the body in these diseases because the mutated protein has other jobs to do in the cell, outside of the lysosome. A protein called cathepsin A, which causes a lysosomal disease called galactosialidosis when mutated, is involved in making elastic fibres. Elastic fibres are very important in making the air sacs in the lungs, which are like little balloons. We are caring for a patient with both galactosialidosis and emphysema and believe the former caused the latter. We can prove our hypothesis by watching under the microscope how well skin cells from the affected patient form elastic fibres. We can also see if they improve their performance by being exposed to various chemicals.

Curing Lafora Disease by Reducing Glycogen Synthesis

Lafora disease (LD) is an incurable fatal genetic disease manifesting as severe constantly worsening teenage-onset epilepsy and is characterized by the presence of abnormal glycogen polymers called polyglucosans, which aggregate into masses called Lafora bodies. These polyglucosans are generated by excessive activity of the enzyme that makes glycogen, glycogen synthase. In our study we will test the role of Everolimus, a drug that inhibits glycogen synthase, and that is in common use for other purposes in other diseases, to cure LD in mice, in preparation for a clinical trial.

Renal Angiomyolipoma and a Constitutional Chromosomal Translocation: Is There a Causative Relationship?

Chromosomes are strings of genes, made out of DNA and protected by proteins. Translocated chromosomes are the result of mis-joining of two different chromosomes after either normal or abnormal breaks in the DNA strands. Depending on what is broken and what is joined together, a disease may or may not follow. Identifying exactly what is present at the joining point has long been a valuable method to find genes involved in specific human diseases. Although, historically, this task has been laborious and expensive, technology has been accelerating this process considerably. Last year we were awarded an RDF grant to try to discover whether any genes are involved in a translocation in a woman with tumors known as renal angiomyolipomas and a translocation between chromosomes 11 and 12. We used the funding to sequence as much DNA as possible around what appeared to be the breakpoints under the microscope-about 6 million base pairs. We have found however that this was not enough. The junction is not contained in those regions. This finding does not make the project any less important to pursue. The question remains for this woman, and for her young daughter who inherited the translocation, whether or not this translocation involves potential cancer genes, either previously known or not known. We now request a second RDF microgrant to continue searching for the junction. This time, however, being one year later in a period of exponential progress in sequencing technology, we will be able to use a much better method for about the same cost: whole genome sequencing.

Meeting the Needs of Children with Skeletal Dysplasias

Children with short statured skeletal dysplasias (conditions affecting bone growth) have particular needs that lead to challenges in everyday life. Our goal is to look at parents' awareness of, and access to, services to help meet these needs so we can provide better care and patient resources.

Proof-of-Principle study: Cofactor Therapy for Weaver Syndrome

We think that adding extra methionine (an amino acid commonly found in a variety of foods, or available as a supplement over-the-counter) to the diet of people with Weaver syndrome will improve their outcome.

Results - In contrast to the enhanced trimethylation activity seen in some somatic mutations that cause leukemia, the WS-associated EZH2 mutants show reduced methylation activity. These results support the hypothesis that WS is caused by loss-of-function mutations in EZH2. Although strategies to reduce EZH2 activity are currently being explored in common cancers such as leukemias, prostate cancer and breast cancer, our data suggest that these strategies may not be effective in rare cancers associated with WS. However, our results suggest that each mutant protein has a different dependence on the concentrations of S-adenosylmethionine (SAM), the methyl donor for the EZH2-mediated methylation of H3K27. Indeed, it does look like increasing the concentration of SAM allows for higher activity of most mutant EZH2 complexes, suggesting that a therapy based on providing extra SAM might be beneficial to some WS patients. Conclusion: Cofactor therapy using S-adenosylmethionine may be effective for some Weaver syndrome patients.

A Small Molecule Cure for GSD Type IV

Glycogen storage disorder (GSD) type IV causes two devastating diseases, a fatal infantile form or adult-onset disease similar to amyotrophic lateral sclerosis (ALS). The disease is characterized by large accumulations of malformed glycogen, also seen in another disease called Lafora disease (LD) and there causes a fatal teenage-onset epilepsy. In a mouse model of LD, the disease was cured by downregulating glycogen synthesis. In this study, we plan to test whether this approach will work for GSD type IV.

Genome-Wide DNA Methylation Profiling in Kabuki Syndrome

Kabuki Syndrome is a condition present at birth affecting many parts of the body and causing intellectual disability. It is also characterized by typical facial features resembling the make-up worn by actors of Kabuki, a Japanese theatrical form. Recently it was discovered that the majority of cases with Kabuki syndrome are caused by mutations in the MLL2 gene, encoding a protein involved in modifications to proteins that package DNA. It is known from model organism studies, that alterations of packaging can affect the state of DNA itself, which is important for defining which regions of DNA are read or expressed. Since DNA modifications can be easily accessed in clinical samples from patients, we suggest that identification of the alterations in DNA modification in the blood of Kabuki patients will further elucidate molecular mechanisms that are disrupted in this disorder. Such data could in future be developed into more cost-effective diagnostic tests, as well as new opportunities for treatment.

Challenges to Talking to Children at Risk for Huntington Disease

Huntington Disease (HD) is an inherited neurodegenerative condition for which no cure exists. This study aims to identify barriers that prevent parents from talking to their children who are at risk for HD. This data will allow us to identify resources to better facilitate parent-child communication.

Characterization of the Carnitine Transporter Gene OCTN 3 in Patients with Peroxisomal Dysfunction

Carnitine is a key vitamin in human energy metabolism that helps transport high energy fats between different compartments of the cell such as peroxisomes, which are important for brain white matter (myelin) formation, and mitochondria, which generate energy. Children with peroxisomal diseases have brain malformations, developmental delay and white matter disease such as adrenoleukodystrophy. Carnitine is transported around the cell by 3 carnitine transporters, one on the cell membrane and two inside. We discovered the third transporter, OCTN3, in human peroxisomes and found 2 children with uncharacterized peroxisomal disease who have reduced or absent OCTN3 protein. Our aim is to identify the OCTN3 gene using the new Next-Generation Sequencing so that we may characterize the mutations in these 2 children as well as others with uncharacterized peroxisomal disease, as these children may respond to high dose oral carnitine therapy.